Functional Delineation of a Protein–Membrane Interaction Hotspot Site on the HIV-1 Neutralizing Antibody 10E8

Antibody engagement with the membrane-proximal external region (MPER) of the envelope glycoprotein (Env) of HIV-1 constitutes a distinctive molecular recognition phenomenon, the full appreciation of which is crucial for understanding the mechanisms that underlie the broad neutralization of the virus. Recognition of the HIV-1 Env antigen seems to depend on two specific features developed by antibodies with MPER specificity: (i) a large cavity at the antigen-binding site that holds the epitope amphipathic helix; and (ii) a membrane-accommodating Fab surface that engages with viral phospholipids. Thus, besides the main Fab-peptide interaction, molecular recognition of MPER depends on semi-specific (electrostatic and hydrophobic) interactions with membranes and, reportedly, on specific binding to the phospholipid head groups. Here, based on available cryo-EM structures of Fab-Env complexes of the anti-MPER antibody 10E8, we sought to delineate the functional antibody-membrane interface using as the defining criterion the neutralization potency and binding affinity improvements induced by Arg substitutions. This rational, Arg-based mutagenesis strategy revealed the position-dependent contribution of electrostatic interactions upon inclusion of Arg-s at the CDR1, CDR2 or FR3 of the Fab light chain. Moreover, the contribution of the most effective Arg-s increased the potency enhancement induced by inclusion of a hydrophobic-at-interface Phe at position 100c of the heavy chain CDR3. In combination, the potency and affinity improvements by Arg residues delineated a protein-membrane interaction site, whose surface and position support a possible mechanism of action for 10E8-induced neutralization. Functional delineation of membrane-interacting patches could open new lines of research to optimize antibodies of therapeutic interest that target integral membrane epitopes.

Publication year: 2022
Authors: Insausti S. 1 2, Garcia-Porras M. 1, Torralba J. 1 2, Morillo I. 1, Ramos-Caballero A. 1, de la Arada I. 1, Apellaniz B. 3, Caaveiro J. 4, Carravilla P. 5, Eggeling C. 5 6 7, Rujas E. 8 9 10, Nieva J. 1 2
  1. Instituto Biofisika (CSIC-UPV/EHU), University of the Basque Country (UPV/EHU), 48080 Bilbao, Spain.
  2. Department of Biochemistry and Molecular Biology, University of the Basque Country (UPV/EHU), 48080 Bilbao, Spain.
  3. Department of Physiology, Faculty of Pharmacy, University of the Basque Country (UPV/EHU), Paseo de la Universidad, 7, 01006 Vitoria-Gasteiz, Spain.
  4. Laboratory of Global Healthcare, School of Pharmaceutical Sciences, Kyushu University, Fukuoka 819-0395, Japan.
  5. Leibniz Institute of Photonic Technology e.V., 07745 Jena, Germany.
  6. Faculty of Physics and Astronomy, Institute of Applied Optics and Biophysics, Friedrich Schiller University Jena, 07743 Jena, Germany.
  7. Medical Research Council Human Immunology Unit, Weatherall Institute of Molecular Medicine, University of Oxford, Oxford OX1 2JD, UK.
  8. Ikerbasque, Basque Foundation for Science, 48013 Bilbao, Spain.
  9. Pharmacokinetic, Nanotechnology and Gene Therapy Group, Faculty of Pharmacy, University of the Basque Country UPV/EHU, 01006 Vitoria-Gasteiz, Spain.
  10. Microbiology, Infectious Disease, Antimicrobial Agents, and Gene Therapy, Bioaraba, 01006 Vitoria-Gasteiz, Spain.
Published in: International Journal of Molecular Sciences, 2022, Vol. 23 p.10767
DOI: 10.3390/ijms231810767


anti-MPER HIV antibody antibody-membrane interactions HIV neutralization MP-SPR lipid-peptide and antibody interaction protein-membrane interaction


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